Bacterial infection is and will continue to be a major health threat, especially with the development of multiple drug resistant pathogens. The success of bacterial infection depends on the ability of bacteria to sense the presence of host cells and produce virulence factors at the right time. Finding a common feature among the diverse array of bacterial sensing sensors can yield effective targets for new antibiotics to fight bacterial infection. In our study of signal transduction required for the invasion of host root hair cells by Sinorhizobium meliloti, we found a bacterial periplasmic protein, ExoR, that appears to regulate the activities of membrane sensors of two-component regulatory systems, which regulate the production of bacterial virulence factors. Our current hypothesis is that ExoR is a global regulator controlling bacterial host cell invasion. ExoR responds to upstream or external signals through the changing of structural conformation or levels of exoR gene expression. The active form of ExoR protein interacts with other membrane sensors such as ExoS and FixL in regulating the expression of genes required for host cell invasion. This hypothesis is developed based on our previous work and the following new findings made in our lab. First, ExoR is a periplasmic protein. Second, ExoR regulates the expression of more than 700 genes. Third, the expression of the exoR gene is regulated by ammonia and the ExoR protein appears to have a site for post-translational modification. Fourth, genetic evidences suggest that ExoR functions directly upstream of the ExoS/Chvl signal transduction pathway. To test the hypothesis, we plan to focus on accomplishing two specific aims. The first is to characterize the signaling mechanism of ExoR by (a) identifying the nodulation stage that requires the function of ExoR;(b) determining the cellular location of ExoR and its glycosaminoglycan modification;and (c) identifying ExoR protein-protein interaction domains. The second is to characterize ExoR-ExoS interactions by (a) characterizing molecular interaction between ExoR and ExoS;(b) identifying regions of ExoR and ExoS that are important for their signaling functions;and (c) determining the global regulatory function of ExoR. This work will provide the first insight for the molecular signaling mechanism of a periplasmic protein and provide a solid base for broad analysis of the global regulatory function of the protein, which will lead to the identification of targets for new classes of antibiotics.